B-Raf Associates with and Activates the NHE1 Isoform of the Na+/H+ Exchanger
Bibliographic record
Abstract
The serine/threonine kinase B-Raf is the second most frequently occurring human oncogene after Ras. Mutations of B-Raf occur with the highest incidences in melanoma, and the most common mutant, V600E, renders B-Raf constitutively active. The sodium proton exchanger isoform 1 (NHE1) is a ubiquitously expressed plasma membrane protein responsible for regulating intracellular pH, cell volume, cell migration, and proliferation. A screen of protein kinases that bind to NHE1 revealed that B-Raf bound to the cytosolic regulatory tail of NHE1. Immunoprecipitation of NHE1 from HeLa and HEK cells confirmed the association of B-Raf with NHE1 in vivo. The expressed and purified C-terminal 182 amino acids of the NHE1 protein were also shown to associate with B-Raf protein in vitro. Because treatment with the kinase inhibitor sorafenib decreased NHE1 activity in HeLa and HEK cells, we examined the role of B-Raf in regulating NHE1 in malignant melanoma cells. Melanoma cells with the B-RafV600E mutation demonstrated increased resting intracellular pH that was dependent on elevated NHE1 activity. NHE1 activity after an acute acid load was also elevated in these cell lines. Moreover, inhibition of B-Raf activity by either sorafenib, PLX4720, or siRNA reduction of B-Raf levels abolished ERK phosphorylation and decreased NHE1 activity. These results demonstrate that B-Raf associates with and stimulates NHE1 activity and that B-RafV600E also increases NHE1 activity that raises intracellular pH. The serine/threonine kinase B-Raf is the second most frequently occurring human oncogene after Ras. Mutations of B-Raf occur with the highest incidences in melanoma, and the most common mutant, V600E, renders B-Raf constitutively active. The sodium proton exchanger isoform 1 (NHE1) is a ubiquitously expressed plasma membrane protein responsible for regulating intracellular pH, cell volume, cell migration, and proliferation. A screen of protein kinases that bind to NHE1 revealed that B-Raf bound to the cytosolic regulatory tail of NHE1. Immunoprecipitation of NHE1 from HeLa and HEK cells confirmed the association of B-Raf with NHE1 in vivo. The expressed and purified C-terminal 182 amino acids of the NHE1 protein were also shown to associate with B-Raf protein in vitro. Because treatment with the kinase inhibitor sorafenib decreased NHE1 activity in HeLa and HEK cells, we examined the role of B-Raf in regulating NHE1 in malignant melanoma cells. Melanoma cells with the B-RafV600E mutation demonstrated increased resting intracellular pH that was dependent on elevated NHE1 activity. NHE1 activity after an acute acid load was also elevated in these cell lines. Moreover, inhibition of B-Raf activity by either sorafenib, PLX4720, or siRNA reduction of B-Raf levels abolished ERK phosphorylation and decreased NHE1 activity. These results demonstrate that B-Raf associates with and stimulates NHE1 activity and that B-RafV600E also increases NHE1 activity that raises intracellular pH.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".